DESCRIPTION: Floodwater mosquitoes, such as Aedes aegypti, oviposit on substrates at the edge of water, and the eggs hatch only after being flooded following adequate rainfall. Therefore, the ability to resist desiccation and other adverse environmental conditions is critical for the survival of these eggs. After oviposition, the chorion hardens in 3-4 hours in a moist environment, and a hardened chorion provides protection for the developing embryo within the chorion. The PI has demonstrated that two interrelated biochemical events, Phenoloxidase/dopa decarboxylase- catalyzed chorion melanization and peroxidase-mediated chorion protein crosslinking are involved in the overall chorion hardening in A. aegypti. Results indicate that the chorion peroxidase-catelyzed chorion protein crosslinking is a vital step and that the peroxidase is significantly different from other described peroxidases and has evolved in a manner that likely reflects its biological role in chorion formation and hardening. The objectives of this grant are to achieve a full understanding of the chorion peroxidase and phenoloxidase in relation to chorion biosynthesis and hardening. Specific aims include 1) isolation of the chorion peroxidase cDNA and assessment of temporal and spatial transcription, 2) production of recombinant protein for enzyme characterization and antibody production, 3) biochemical characterization of chorion peroxidase, and 4) the biochemical and molecular characterization of chorion phenoloxidase. Mosquito-borne pathogens continue to have a major impact on human health throughout the world, and the development of drug resistant parasites and insecticide resistant mosquito vectors urgently demands that new mosquito or parasite control strategies be developed. Disruption of the required physio-biochemical processes that are necessary for the normal development of mosquitoes is one approach for the control of mosquito-borne disease. A better knowledge of the physiology and biochemistry of chorion formation in mosquitoes is critical to a thorough understanding of reproductive processes in this important group of disease vectors. Survival of eggs for extended periods of time in adverse-environmental conditions, as is required for floodwater mosquitoes, is critically dependent on the biochemical process of chorion biosynthesis and hardening. It is anticipated that an elucidation of the biochemical pathways and their regulation required for egg chorion development might provide new approaches for the control of mosquitoes through the disruption of required reproductive processes.